健康与遗传心理学研究室知识库

越来越多的证据显示神经和免疫存在交互作用，它们共同调节脑的结构和功能的成熟化过程。在这一过程中，脑内主要免疫细胞一小胶质细胞及一类主要由其分泌的免疫因子一肿瘤坏死因子a C tumor necrosis factor alpha, TNFa)的表达、分布和功能在出生后呈现出动态发展变化，对神经环路的形成、维系和重塑十分重要。早期应激可以通过干扰中枢免疫的“程序化”发展进程对脑结构和功能造成长期影响，增加后期精神疾病的患病风险。认知灵活性是一种由前额叶皮质介导的高级脑功能，其损伤是多种常见精神疾病的主要症状，也是其发病、预后和药物治疗反应的重要风险因素。我们前期的研究运用青少期社会挫败应激，一种拟人类欺侮的病因学应激范式，稳定诱发了成年(而非青少期)小鼠认知灵活性损伤，主要表现为内侧前额叶皮质(medial prefrontal cortex,mPFC)介导的外维度定势转换能力受损。采用这一模型，本研究探讨青少期社会应激诱导的行为介导脑区mPFC小胶质细胞和TNFa的动态改变与上述认知功能障碍发生发展的关系，为建立基于免疫调节的认知功能障碍防治策略提供新的研究范式和实验证据。

首先，本研究描绘了青少期社会应激诱导的mPFC小胶质细胞和TNFa表达的变化过程;然后，从成年期治疗和青少期早期干预两个方面分别考察了免疫改变与认知灵活性损伤发生发展之间的关系。采用注意定势转移任务测试动物的认知灵活性水平;采用IHC-Fr和激光扫描共聚焦显微镜技术测量小胶质细胞的数量、形态和活化状态;采用Western blot技术检测小胶质细胞特异性结构标记分子一离子钙结合衔接分子1 Lionized calcium binding adaptor molecule 1,Iba1)和活化标记分子一分化抗原簇l 1b C cluster of differentiation molecule l11b,CD1lb)，以及TNF。的蛋白表达水平;采用实时荧光定量PCR技术检测TNFa,的mRNA表达水平。

结果显示:1)在行为水平上，与前期的研究发现一致，青少期社会应激诱发了成年(而非青少期)小鼠由mPFC介导的外维度定势转换能力损伤。在分子水平上，与对照组相比，应激组小鼠mPFC Ibal和TNF。的蛋白表达水平呈现出类似的变化过程，表现为在应激期间短暂升高，随后逐渐降低，至成年期则明显降低，提示青少期应激诱发免疫反应短时增强到长时抑制的变化过程。2)对免疫反应长时抑制的功能分析结果显示，青少期社会应激诱发了成年小鼠mPFC小胶质细胞数量减少、一级突起长度缩短等结构和形态学改变，以及TNFa, mRNA和蛋白表达水平降低，提示经历青少期应激的成年小鼠mPFC小胶质细胞和TNF。功能不足。3)成年期慢性单胺氧化酶抑制剂反苯环丙胺C Tranylcypromine, TCP)治疗能够改善青少期社会应激诱导的成年小鼠外维度定势转换能力损伤，并恢复mPFC降低的Ibal和TNFa蛋白表达水平。4)成年期单次急性脂多糖CLipopolysaccharide, LPS)免疫激活能够快速改善青少期社会应激诱导的成年小鼠上述认知功能和免疫分子改变。_5 )通过在青少期应激期间给与小胶质细胞抑制剂米诺环素拮抗应激诱导的免疫激活反应，可以预防成年期外维度定势转换能力的损伤以及mPFC Ibal和TNFa蛋白表达水平的降低。

本研究结果表明，mPFC小胶质细胞和TNFa动态改变参与青少期应激诱导的认知灵活性损伤的发生发展，免疫调节具有精神疾病相关认知功能障碍快速治疗和早期预防的潜力。

Increasing evidence shows that neuro-immune interactions coordinately regulate the maturation of brain structure and function. During this process, the expression, distribution and function of microglia, the primary immune cells of the brain, and tumor necrosis factor alpha (TNFα), an immune molecule mainly released by microglia, develop dynamically after birth and play crucial roles in the formation, maintenance and remodeling of neuronal circuits. Early life stress can cause long-term effects on brain structure and function through disturbing the "programmed" development of brain immunity, thus increase psychopathological risk in the future. The impairment of cognitive flexibility, an executive function mediated by prefrontal cortex, is increasingly considered as a main symptom and as an important risk factor for the pathogenesis, prognosis and drug response of several common psychiatric disorders. Our previous studies demonstrated that adolescent social defeat stress, an etiological stress model analogous to human bullying, induced impairment of cognitive flexibility, characterized by deficit of extra-dimensional shifting ability mediated by the medial prefrontal cortex (mPFC) in adult, but not adolescent mice. Using this model, the present study aimed to investigate the relationship of dynamic changes of microglia and TNFa in this brain region and the development of cognitive dysfunction, with purposes to provide new research paradigm and experimental evidence to set up preventive and therapeutic strategies for cognitive dysfunction based on immune modulation.

Firstly, this study profiled the time courses of microglia and TNFα changes in the mPFC induced by adolescent social stress. Then, the relationship of the immune alterations and the development of cognitive deficit induced by adolescent social stress was examined from two aspects, symptomatic treatment during adulthood and preventive intervention during early adolescence. The attentional set-shifting task was performed to assess cognitive flexibility; the IHC-Fr and laser scanning confocal microscopy were conducted to analyze the number, morphology and activation state of microglial cells; the Western blot method was conducted to detect the protein levels of ionized calcium binding adaptor molecule 1 (Iba1)，a constitutively expressed marker specific to microglia, and cluster of differentiation molecule 11b (CDllb), a marker for microglial activation, as well as TNFα; the qRT-PCR method was conducted to detect the mRNA level of TNFα.

Results showed that: 1) At behavioral level, consistent with our previous findings, adolescent social stress induced deficit of extra-dimensional shifting ability in adult, but not adolescent mice. At molecular level, compared to the controls, stress-exposed animals showed parallel variations in the protein expression of Ibal and TNFa in the mPFC, with a transient increase during stress exposure, then a gradual decrease after the end of stress, leading to a significantly decreased level in adulthood. These suggested that adolescent social stress induced dynamic alterations of immune response from short-term enhancement to long-term suppression. 2) Further functional analysis of the above long-term suppression revealed that adolescent social stress induced structural and morphological changes of microglial cells, including decreased numbers and reduced primary process lengths, as well as reduced mRNA and protein expression of TNFa in the mPFC of adult mice, suggesting deficiency of microglia and TNFa in the mPFC of mice exposed to adolescent stress. 3) Chronic treatment with monoamine oxidase inhibitor Tranylcypromine (TCP) during adulthood reversed the deficit of extra-dimensional shifting ability and the decrease of Ibal and TNFa protein expression in the mPFC of mice exposed to adolescent social stress. 4) Single acute treatment with immune activator Lipopolysaccharide (LPS) during adulthood rapidly reversed the above-mentioned alterations in cognitive function and immune molecules induced by adolescent social stress. 5) Adolescent intervention with immune inhibitor minocycline during stress exposure to block initial immune activation induced by dolescent social stress prevented the subsequent development of cognitive deficit and immune alterations in the mPFC of adult mice.

These results indicated that the turbulence of microglia and TNFa in the mPFC induced by adolescent stress may participate in the development of cognitive flexibility impairment. Our results provided a potentially new avenue for rapid treatment and early prevention of cognitive deficits in psychiatric disorders based on immune modulation.